Mechanoresponsive Engrailed-1-negative fibroblasts activate Engrailed-1 to promote fibrosis in wound healing

7.项目摘要/摘要 成年人的皮肤通过形成纤维性疤痕组织来愈合，这可能会导致毁灭性的毁容、生长 限制和永久性功能丧失。尽管有太多的临床选择，但目前还没有治疗策略 成功地阻止或逆转这种纤维化过程，疤痕及其后遗症使美国付出了 每年200亿美元。新疗法的开发进展受到了严重阻碍 对导致疤痕形成的特定细胞群缺乏了解。2015年，我们小组报告说 增强型-1(EN-1)系阳性成纤维细胞(EPF)是背部瘢痕的主要来源 在出生后的小鼠中产生。在胎儿怀孕早期，小鼠通过皮肤再生无疤痕愈合，这是一个理想的结果 由EN-1系阴性的成纤维细胞(ENF；主要的胎儿成纤维细胞)介导。然而，它并没有 确定ENF是否有助于出生后伤口愈合。在这项提案中，我们首次探索了 创面内ENF向促纤维化EPF(生后衍生EPF；pEPF)的生后转化 环境。首先，在一种新的转基因小鼠模型中，组织学、免疫组织化学和创伤 用于研究创伤愈合过程中ENF向pEPF的转化。通过检查ENF的行为 伤口环境中的亚群(来源于乳头状真皮、网状真皮和真皮下组织) 在他莫昔芬诱导的EN-1激活的小鼠模型中，我们将准确地定义ENF，以证实我们的发现 导致促纤维化的pEPF的人群。第二，我们将建立特定的创伤环境线索 推动了从ENF到EPF的转变。考虑到机械力既可以调节SCAR负担，也可以调节SCAR负担 我们将使用体外和体内模型来检测机械环境对成纤维细胞活性的影响 EN-1激活。我们将进一步使用转录和表观基因组图谱来探索 ENF向EPF转化过程中的机械转导信号和pEPF功能。第三，建立了一个 创伤ENF中EN-1激活的机械转导机制，我们将抑制 机械转导信号，目的是阻止ENF到EPF的转变。具体地说，我们将评估 阻断机械转导是否导致ENF介导的伤口愈合和减少纤维化。我们的 最终的翻译目标是开发靶向成纤维细胞以促进再生的治疗方法 治愈。总的来说，拟议的工作将显著增强我们对关键分子和 皮肤疤痕形成的细胞决定因素，提示新的抗疤痕疗法的发展，并脱落 皮肤瘢痕成纤维细胞的细胞起源。

7. Project Summary/Abstract Adult human skin heals by developing fibrotic scar tissue, which can result in devastating disfigurement, growth restriction, and permanent functional loss. Despite a plethora of clinical options, no current treatment strategies successfully prevent or reverse this fibrotic process, and scars and their sequelae cost the United States over $20 billion every year. Progress towards the development of new therapies has been significantly hindered by a lack of understanding of the specific cell populations responsible for scarring. In 2015, our group reported that Engrailed-1 (En-1) lineage-positive fibroblasts (EPFs) are responsible for the vast majority of dorsal scar production in postnatal mice. In early fetal gestation, mice heal scarlessly via skin regeneration, an ideal outcome mediated by En-1 lineage-negative fibroblasts (ENFs; the predominant fetal fibroblast). However, it has not been established if ENFs contribute to postnatal wound healing. In this proposal, we explore for the first time the postnatal conversion of ENFs to pro-fibrotic EPFs (postnatally-derived EPFs; pEPFs) within the wound environment. First, histology, immunohistochemistry, and wounding in a novel transgenic mouse model will be used to study the conversion of ENFs to pEPFs during wound healing. By examining the behavior of ENF subpopulations (derived from papillary dermis, reticular dermis, and hypodermis) in the wound environment and confirming our findings in a tamoxifen-inducible mouse model of En-1 activation, we will precisely define the ENF population that gives rise to pro-fibrotic pEPFs. Second, we will establish the specific wound environment cues that drive ENF-to-EPF transition. Given that mechanical forces are known to modulate both scar burden and fibroblast activity, we will use in vitro and in vivo models to examine the effects of mechanical environment on En-1 activation. We will further use transcriptomic and epigenomic profiling to explore the role of mechanotransduction signaling in ENF-to-EPF transition and pEPF function. Third, having established a mechanotransduction mechanism underlying En-1 activation in wound ENFs, we will inhibit mechanotransduction signaling with the goal of blocking ENF-to-EPF transition. Specifically, we will assess whether blocking mechanotransduction results in ENF-mediated wound healing with reduced fibrosis. Our ultimate translational goal is to develop therapeutics that target fibrogenic fibroblasts to promote regenerative healing. Collectively, the proposed work will significantly enhance our understanding of the key molecular and cellular determinants of cutaneous scarring, inform the development of novel anti-scarring therapies, and shed light on the cellular origin of dermal scarring fibroblasts.